Antimicrobial drugs, particularly those used in animal feed or to treat animals, have been found in milk and food products, such as for example, beta lactams like penicillin and/or tetracyclines, and more recently sulfa drugs like sulfonamides, such as sulfamethazine, have been detected in food products. The detection of these drugs at low levels are most important, particularly sulfa drugs which are allergenic and possibly carcinogenic in nature, even at low levels. Milk is routinely tested for the presence of drugs, like beta lactam, by the use of an inhibition disc assay method. However, such assay method does not detect sulfa drugs or certain other antimicrobial drugs at very low levels.
One technique for detecting sulfa drugs in milk at low levels is by the employment of high pressure liquid chromatography (HPLC) or by receptor assay test method (see U.S. Pat. Nos. 4,239,745 and 4,239,852, both issued Dec. 16, 1980). Both of such test methods are cumbersome and require costly equipment and highly trained technicians.
A microbiological growth inhibition test method to detect sulfonamides in milk for example has been reported and described in European patent specification 79200277.6 by Beukers et al, published Dec. 12, 1979. This test method has been described as an improvement on the test described in British patent specification 1 467 439, published Mar. 16, 1977 to Lameris et at. The Beukers et at test method places spores of Bacillus stearothermophilus (BST) into a buffered agar solution, and the agar is solidified to form a support medium. The BST in the agar solution is prevented from germinating by lack of nutrients in the agar and/or by low temperature. The nutrients for the growth of the BST are prepared separately and are either placed in a tablet or on a disc, with the tablet or disc placed on the solid agar medium containing the BST before carrying out a test. A test sample is then added and the sensitivity for sulfa drugs is increased by the addition of trimethoprin while an indicator is added either to the agar solution prior to solidification or to the nutrients so as to provide for increased sensitivity for sulfa drugs. However, sensitivity is not less than about 100-500 ppb.
It is desirable to provide for a microbial inhibition test kit, method and apparatus which is simple and effective to employ and is sufficiently sensitive to detect very low levels of drugs, particularly sulfamethazine in milk and other products.